Since the invention of microtechnology for realizing integrated semiconductor structures for microelectronic chips in the 1950s, these lithography-based technologies have been applied to a wide variety of applications ranging from entertainment (for example gaming consoles, MP3 players etc), through consumer electronics (digital cameras, personal computers, personal data assistants (PDAs) etc), to advanced avionics and telecommunications. Bolstered in the 1960s by generally CMOS compatible micrometer or sub-micrometer sized mechanical structures, known commonly as Micro Electro Mechanical Systems (MEMS), such integrated semiconductor structures have allowed for pressure sensors, airbag sensors, tunable capacitors, inductors and resonators, pivotable mirrors, switches, valves, pumps as well as other mechanically movable structures to become common elements of many consumer and high volume applications.
Concurrently such advances in integrated semiconductor devices also triggered advances in printed circuit boards as interconnections and assemblies became denser, faster, three-dimensional, wire-bondable, solder reflow compatible, and addressed heat management. Accordingly synthetic resin bonded paper materials such as FR-2 were replaced with UV stabilized tetrafunctional epoxy resins, such as FR-4, and ceramics, e.g. aluminum oxide and aluminum nitride, co-fired ceramic green sheets, ceramic packages with copper tungsten inserts etc. Advances were also made in exploiting silicon and semiconductor materials in the microwave domains as well as the photonic domain.
By appropriate combinations of these technologies engineers and material scientists developed solutions to begin replacing bulky, expensive discrete test, evaluation and measurement structures with compact, low cost, replacements that could put tens, hundreds, even thousands of measurand sites within the same footprint. With the advent of fluid based micro-treatments for analysis of biological specimens (so-called μTAS) such systems became feasible for detecting and characterising samples, exploiting techniques such as capillary electrophoresis, chromatographic separation, DNA microarrays, and physiochemical changes of proteins. Coincident requirements for testing within biological and bio-chemical applications such as within the environmental and pollution monitoring, chemical analysis, medical diagnostics and cellomics, together with synthetic chemistry applications involving rapid screening and microreactors for pharmaceutics have also established demand for low cost measurement solutions and high numbers of measurements to be made rapidly.
Also driving these developments has been the potential to fabricate test arrays for these diverse applications within a silicon platform, which in different forms such as native silicon, micro- and macro-porous silicon, and nitrocellulose-coated variants offers potential for low cost manufacturing by leveraging existing high volume semiconductor manufacturing techniques, high biocompatibility allowing prolonged use rather than discrete measurements, and potential integration of microfluidics, sensors, characterization and analysis elements within circuits integrating CMOS electronics. Even within less advanced applications the advances in printed circuits, ceramic substrates, etc allow for low cost arrays to be provided with tens, hundreds to thousands of test sites.
Amongst the benefits of these different manufacturing approaches are:                ability to characterise samples with low fluid volumes which means less waste, lower reagents costs and less required sample volumes for diagnostics;        faster analysis and response times due to short diffusion distances, fast heating, high surface to volume ratios, small heat capacities, etc;        better process control because of a faster response of the system, e.g. thermal control for exothermic chemical reactions;        compactness of the systems due to integration of much functionality and small volumes;        massive parallelization due to compactness, which allows high-throughput analysis and multiple analysis processes within a single integrated circuit;        lower fabrication costs, allowing cost-effective disposable chips, fabricated in mass production, and wide-spread deployment;        safer platform for chemical, radioactive or biological studies because of integration of functionality, smaller fluid volumes and stored energies.        
When considering any system intended to measure, characterise, analyse or evaluate a particular attribute then the system would normally be considered to be composed essentially of two parts, the transducer which generates a variation in an electrical characteristic in dependence of the measurand, and the measurement electronics which receive and convert the transducer output to a measured value for the measurand. This electrical characteristic may for example be resistance but it is more likely to be a variation in inductance, capacitance, resonant frequency of an oscillator, etc either in isolation or in conjunction with others including resistance. However, in some applications such as bio sensors then the system is best considered to be comprised of three parts:
the sensitive biological element, which may be biological material, e.g. tissue, microorganisms, organelles, cell receptors, enzymes, antibodies, nucleic acids, etc, or a biologically derived material or biomimic, wherein the sensitive elements can be created by biological engineering;
the transducer or the detector element, which works in a physicochemical way; optical, piezoelectric, electrochemical, etc., that transforms the signal resulting from the interaction of the analyte with the biological element into another signal (i.e., transducers) that can be more easily measured and quantified; and
the associated electronics or signal processors that are primarily responsible for the display of the results in a user-friendly way.
As noted supra typically the transducers will present a variation in impedance rather than a simple change in resistance to the electronics and signal processors, and as such the effective electrical circuits these transducers present will have energy storage and dissipation properties which will vary with applied frequency of a probe electrical signal, i.e. their AC properties. Accordingly over the past few years the approach of electrochemical impedance spectroscopy (EIS), also referred to as dielectric spectroscopy or impedance spectroscopy, has grown tremendously and is deployed in a wide variety of scientific fields such as fuel cell testing, biomolecular interaction, micro structural characterization, and electrochemical systems. EIS measures the impedance of a system over a range of frequencies allowing variations in the real and imaginary components to be determined as well as variations in the phase relationship of the output signal with respect to the input excitation signal. Additionally, EIS reveals information about reaction mechanisms within electrochemical processes as different reaction steps will dominate at certain frequencies, and the analysis of the frequency response obtained by EIS can help identify these processes as well as determine rate limiting steps.
However, an issue with EIS systems, and the electronics/signal processors within analysis systems generally is that systems which either perform multiple measurements for a single measurand in order to obtain position dependent information or perform analysis of multiple measurands for multiple samples or even single measurands on multiple samples is the third critical element, the associated electronics. For example, each transducer or detector element there is required an associated analog-to-digital converter (ADC) to convert the analog output of the transducer or detector element to a digital representation that can be read by subsequent digital processing circuitry or microprocessor to provide the result of the measurement made using the transducer or detector element. This requirement is exacerbated further when considering deployment of such analysis systems in environments other than as laboratory test equipment in that resolution of the measurement is determined by the number of bits of the ADC, and typically ADCs with a large number of bits are expensive devices. Equally, fast ADCs allowing the measurements to be made dynamically are similarly expensive devices.
However, in many instances the voltage levels required by ADCs are of the order of a few volts which may affect the biosensor and thereby affect the measurement itself. As a result electrochemical impedance measurements typically require that the voltages applied to the biosensor be of order 5 millivolts (5 mV) to 50 millivolts (50 mV) and may vary in frequency, for example over a range of 1 milliHertz (0.001 Hz or 1 mHz) to 1 MegaHertz (1 MHz), according to the measurement being performed and the sensor employed.
Unfortunately at present like high resolution, fast ADCs systems with low signal levels are typically very expensive as well as being large, heavy laboratory based instrumentation. In many instances these are developed around a frequency response analyser (FRA), such as those shown for example in FIG. 1 including the “Alpha-A” high performance modular measurement system 110 from Novocontrol, “Reference 600” Potentiometer/Galvanometer FRA 120 from Gamry Instruments, “LEIS370” Localized Electrochemical Impedance System 130 from Princeton Applied Research, “1255A Frequency Response Analyser” 140 from Solartron Analytical, “Model 3120” FRA 150 from Venable Instruments, “Model 2505” FRA 160 from Clarke-Hess Communications Research, and the “RA Series 01” FRA 170 and “SA Series 01” modular FRA 180 from Core Technology Group.
Referring to FIG. 2 there are shown commercial EIS systems targeted to biotechnology applications, these being “ECIS Z” 210 from Applied Biophysics and the “96X” series analyzer 220 from ACEA Biosciences. Hence, it is evident that whilst semiconductor manufacturing processes and biochemical processes can provide low cost assay elements, ranging from implantable glucose monitoring structures through to very large disposable assay trays the benefits of EIS at present are limited to environments to such as laboratories, medical clinics, etc where the deployment of such large, expensive systems can be justified or permits their use. Additionally such systems typical present significant limitations in their use through the requirements for calibration.
Hence, it would therefore be beneficial to provide a compact, fast (i.e. high-throughput) EIS electrochemical impedance spectrometry system (FSCEISS) that is self-calibrating. It would be further beneficial if the FSCEISS was implementable with electronics and software/firmware that supported implementations in multiple technologies. For example, it would be beneficial if ultimately the FSCEISS could be implemented as a single monolithic integrated circuit to fully leverage CMOS silicon electronics for very high volume low cost applications such as blood glucose monitoring for diabetics and/or insulin dosage control for type 1 diabetics. The World Health Organization projects that the number of diabetics requiring regular periodic monitoring will exceed 350 million by 2030 and of these up to 50 million will be Type 1 diabetics requiring continuous closed loop delivery systems to control their insulin levels.
Alternatively, in other applications, such as medical clinics, environmental monitoring stations, biochemical monitoring etc it would be beneficial for the FSCEISS to be manufactured leveraging for example with hybrid electronic integration using multi-chip modules (MCMs) or packaged integrated circuits with PCB assembly techniques. Such an FSCEISS thereby allows for an implementation to be tailored to the cost—volume—performance tradeoffs of the particular application.
Another aspect of EIS measurements systems is the excitation signal, which as noted supra may for example be within the range 1 milliHertz (0.001 Hz or 1 mHz) to 100 kilohertz and have an amplitude between 5 mV to 50 mV. Providing a source covering 8 orders of magnitude in frequency and low stable output voltage is another challenging aspect for electronics, suited generally to the large, laboratory style instruments described supra in respect of FIGS. 1 and 2. Commercial synthesizers or digital-to-analog converters (DACs) such as Analog Devices AD766 16 bit 390 kS/s DAC operating at ±3V are capable of achieving such output amplitudes although with a resolution of 0.05 mV at the lowest range limit this signal is generated with the equivalent of a 6-bit ADC. Accordingly the excitation signal is not a high purity single frequency. It would therefore be beneficial to provide a method of analysis that accounted for an imperfect excitation signal.
It is accordingly an intention of this invention to provide a high throughput self-calibrating electrochemical impedance spectroscopy measurement system (FSCEISS) that is compatible with a variety of measurement environments by providing low footprint, high speed, broad frequency response, and an ability to operate with a significant number of measurement sites, the significant number of measurement sites representing measurements that are either spatially distributed and/or multiple biochemical species.
It is also an intention of the invention for such high throughput self-calibrating electrochemical impedance spectroscopy measurement systems to be implementable in a manner that is low cost and permits implementation in different electronic format, ranging from monolithic integration, hybrid integration, through to discretes according to the market dynamics the FSCEISS addresses and provides self-calibration allowing long term use associated with a single user or within environments such as medical clinics where appropriate equipments and expertise is not available.